Cytokinin-Mediated Translational Control of Protein Synthesis in Cultured Cells ofGlycinemax

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Polyribosome formation was stimulated by cytokinin treatment of cultured cells of Glycine max cv. Funk Delicious. When suspension cultures were given 0-5 pM zeatin after 24 h in culture in medium lacking a cytokinin, a nearly 2-fold increase in the polyribosome/monoribosome ratio occurred over the subsequent 3 h. The effect of actinomycin D and of 5-fluorouridine on RNA synthesis and on the polyribosome/monoribosome ratios of these cells was examined. Actinomycin D at 5 and 20 pg/ml_1inhibited total RNA synthesis by 39 and 60%, respectively, as measured by [3H]uridine incorporation into acid-precipitable material. The degree of inhibition of precursor incorporation into polyribosomal RNA was similar. At 0-1 mM, 5 fluorouridine inhibited [3H]uridine incorporation by 76%, and [3H]guanosine incorporation by 66% into polyribosomal RNA after 3 h of treatment. Fractionation of the polyribosomal RNA by oligo(dT)-cellulose chromatography demonstrated that low concentrations of both actinomycin D (5 pg ml-1) and 5-fluorouridine (0-1 mM) inhibited the synthesis of ribosomal RNA to a greater extent than the poly(A)-containing fraction of the messenger RNA. Synthesis of the poly (A) -containing RNA was inhibited by 24% with 5 pg ml-1 actinomycin D and by 30% with 0-1 mM 5-fluorouridine. At the above concentrations, these two inhibitors reduced the polyribosome/monoribosome ratio of the cytokinin-deprived cells over a 3 h period, but they did not prevent cytokinin-induced polyribosome formation. These results provide further evidence that cytokinin regulates polyribosome levels through an effect on protein synthesis at the translational level